New application of traditional Chinese medicine composition

Abstract

The invention provides a new application of a traditional Chinese medicine composition, the traditional Chinese medicine composition is composed of salvianolic acid B and senkyunolide I. The synergistic effect of salvianolic acid B and senkyunolide I is utilized, neuroinflammation is inhibited through multi-target regulation, a blood-brain barrier structure is maintained, brain injury is reduced, a safe and effective treatment strategy is provided for cerebral arterial thrombosis, and the traditional Chinese medicine composition has a wide application prospect. The compound can be used for preparing a blood-brain barrier protective agent and a medicine for preventing or treating cerebral arterial thrombosis, and has an important clinical application value.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical applications, and in particular to a new use of a traditional Chinese medicine composition. Background Art

[0002] Ischemic stroke is the most common type of stroke, accounting for more than 80% of the stroke incidence. Its characteristics are the disruption of the blood-brain barrier, leading to brain tissue edema, inflammatory response, and nerve function damage. The blood-brain barrier is a physical and biochemical barrier between the central nervous system and the peripheral circulation, used to maintain the homeostasis in the brain. After ischemic injury, the disruption of the blood-brain barrier will lead to aggravated brain tissue damage. Therefore, protecting the blood-brain barrier is one of the important strategies for the treatment of ischemic stroke.

[0003] Existing clinical drugs such as tissue plasminogen activator (tPA) are limited in application due to the narrow treatment time window and the possible aggravation of blood-brain barrier damage. Therefore, developing new drugs for protecting the blood-brain barrier has important clinical significance. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a new use of a traditional Chinese medicine composition.

[0005] The technical solution adopted by the present invention is:

[0006] Use of a traditional Chinese medicine composition in the preparation of a blood-brain barrier protector, wherein the traditional Chinese medicine composition is composed of salvianolic acid B and ligustilide I.

[0007] Preferably, in the above use of the traditional Chinese medicine composition, the weight ratio of salvianolic acid B to ligustilide I is 1:2.6 - 3.

[0008] Preferably, in the above use of the traditional Chinese medicine composition, the cell usage amounts of salvianolic acid B and ligustilide I are 5.7 μMole and 1 μMole respectively.

[0009] The above traditional Chinese medicine composition protects the blood-brain barrier through the following mechanisms:

[0010] 1. Inhibiting the increase in blood-brain barrier permeability caused by ischemia-reperfusion injury;

[0011] 2. Upregulating the expression of tight junction proteins ZO-1 and Occludin;

[0012] 3. Blocking the neuroinflammatory response mediated by the TOLL-like receptor 4 / NF-κB / matrix metalloproteinase 9 signaling pathway.

[0013] Preferably, in the above use of the traditional Chinese medicine composition, the blood-brain barrier protector is a drug for preventing or treating ischemic stroke.

[0014] Preferably, for the use of the above traditional Chinese medicine composition, the traditional Chinese medicine composition is prepared by the following method:

[0015] (1) Weigh the monomer standard products of ligustilide I and salvianolic acid B respectively according to the amount.

[0016] (2) After dissolving the monomer standard products of ligustilide I and salvianolic acid B in methanol respectively, dilute and mix them with pure water.

[0017] A blood-brain barrier protector, comprising the above traditional Chinese medicine composition and pharmaceutically acceptable excipients.

[0018] The beneficial effects of the present invention are:

[0019] The traditional Chinese medicine composition is composed of salvianolic acid B and ligustilide I. By utilizing the synergistic effect of salvianolic acid B and ligustilide I, it inhibits neuroinflammation and maintains the blood-brain barrier structure through multi-target regulation, reduces brain damage, provides a safe and effective treatment strategy for ischemic stroke, can be used to prepare blood-brain barrier protectors and drugs for preventing or treating ischemic stroke, and has important clinical application value. Description of the Drawings

[0020] Figure 1 For the fingerprint maps and methodological investigations of salvianolic acid B and ligustilide I;

[0021] Figure 2 For the effects of salvianolic acid B and ligustilide I on the permeability of the 3D blood-brain barrier model;

[0022] Figure 3 For the expression changes of tight junction proteins ZO-1 and Occludin;

[0023] Figure 4 For the regulation analysis of the TOLL-like receptor 4 / NF-κB / matrix metalloproteinase 9 signaling pathway;

[0024] Figure 5 For the molecular docking and kinetic simulation results of salvianolic acid B and ligustilide I with matrix metalloproteinase 9; [[ID=�9]]

[0025] Figure 6 For the changes in the TOLL-like receptor 4-NF-κB-matrix metalloproteinase 9 signaling pathway in the 3D blood-brain barrier model; [[ID=^3]]

[0026] Figure 7 For the evaluation of blood-brain barrier permeability, cerebral infarction volume and neurological function in the mouse model;

[0027] Figure 8 For the in vivo expression analysis of the proteins in the TOLL-like receptor 4 / NF-κB / matrix metalloproteinase 9 axis. Detailed Embodiments

[0028] To enable those skilled in the art to better understand the technical solution of the present invention, the following further elaborates on the technical solution of the present invention in conjunction with the accompanying drawings and specific embodiments.

[0029] In the following examples, the experimental materials and methods used are conventional materials and methods unless otherwise specified. The terms used have the general meanings known to those skilled in the art if not clearly stated or defined.

[0030] The materials used include: human astrocytes (HA), human brain microvascular endothelial cells (HBMEC), human primary cerebral vascular pericytes (HBVP). High-content screening system, microcentrifuge, floor-standing refrigerated centrifuge (RC6, Thermo Company, USA), polylysine, agar, fluorescent dye FITC-labeled dextran (FITC Dextran 70KDa and 10KDa), Hoechst33342, and calcein AM / propidium iodide, sugar-free DMEM medium, diaphragm vacuum pump (GM-0.33A, Shanghai Shengke Instrument Equipment Co., Ltd.), 96-well U-shaped plates, hypoxia chamber, GraphPad Prism 7 statistical software package.

[0031] Example 1

[0032] A traditional Chinese medicine composition is composed of salvianolic acid B and senkyunolide I and is prepared by the following method:

[0033] Weigh salvianic acid, protocatechuic acid, protocatechuic aldehyde, chlorogenic acid, caffeic acid, ferulic acid, senkyunolide I, senkyunolide H, salvianolic acid B, and salvianolic acid A (0.325 mg, 0.500 mg, 0.545 mg, 0.250 mg, 0.600 mg, 0.405 mg, 0.580 mg, 0.395 mg, 0.540 mg, 0.570 mg) respectively and place them in a 20 mL volumetric flask to prepare a mixed standard solution. Add methanol to the scale line for the establishment and identification of the HPLC fingerprint of the salvia-chuanxiong sample. At the same time, accurately weigh 1.585 mg and 0.6 mg of senkyunolide I and salvianolic acid B standards respectively, dissolve them in 1 ml of methanol, prepare standard solutions with mother liquor concentrations of 1.585 mg / mL and 0.6 mg / mL respectively, and draw concentration curves by stepwise dilution to quantitatively analyze the contents of these two components in salvia-chuanxiong, providing guidance and reference for subsequent drug administration.

[0034] Prepare the Salvia miltiorrhiza - Ligusticum chuanxiong sample. Take 1 mL of the originally stored liquid from the ampoule, add 5 mL of a mixture of methanol and water (50:50, v:v), stir for 3 minutes, vortex for 5 minutes, and then sonicate at 100 Hz and 35 °C for 15 minutes. Remove the sample and centrifuge at 12000 rpm / min for 5 minutes. Then filter the supernatant through a 0.22 - micron nylon membrane to obtain the sample solution.

[0035] Separate the components in Salvia miltiorrhiza - Ligusticum chuanxiong using an Agilent 1290 Infinity II high - performance liquid chromatograph equipped with a diode - array detector. Collect the HPLC fingerprint of the above - prepared test solution. The chromatographic column is Waters ACQUITY UPLC HSS T3 (2.1×100 mm, 1.8 μm), the mobile phase is 0.1% formic acid / water (A) and acetonitrile (B), and the gradient elution program is as follows: 0 - 15 min, 5 - 14% B; 15 - 20 min, 14 - 17% B; 20 - 26 min, 17 - 23% B; 26 - 27 min, 23 - 25% B, 27 - 31 min, 25% - 27% B, 31 - 34 min, 27% - 30% B, 34 - 40 min, 30% - 90% B, 40 - 43 min, 90% - 5% B, 43 - 45 min, 5% B. The flow rate is maintained at 0.3 mL / min, and the injection volume is 2 μL. The detection wavelengths are 254, 280, and 320 nm, and the column temperature is 35 °C.

[0036] Use OpenLab CDS software for system control and data analysis. The precision, stability, linearity, and repeatability of the method are verified. The precision, stability, and reproducibility of the chromatographic method are tested by analyzing a six - replicate sample. The linearity of the chromatographic method is determined using a standard solution.

[0037] Based on fingerprint research, determine senkyunolide I and salvianolic acid B, which are the main components of Ligusticum chuanxiong and Salvia miltiorrhiza respectively, as the research objects for subsequent drug administration. The results are as Figure 1 shown. A comprehensive methodological investigation for quantitative analysis of the compound shows that the maximum ultraviolet absorption wavelength of senkyunolide I is 280 nm, while that of salvianolic acid B is 320 nm. Figure 1 B Based on the standard solution of the prepared stock solution, gradually dilute to draw a concentration curve. Determine the ratio of senkyunolide I to salvianolic acid B in Salvia miltiorrhiza - Ligusticum chuanxiong to be approximately 1:2.6 - 1:3 through multiple measurements and averaging. Determine the administration concentrations of senkyunolide I and salvianolic acid B in cells to be 1.67 μg / ml (5.7 μM) and 0.72 μg / ml (1 μM) based on the concentrations in the Salvia miltiorrhiza - Ligusticum chuanxiong sample ( Figure 1B). Therefore, the subsequent cell administration dose was dissolved in PBS to be 5.7 uMol ligustilide I and 1 uMol salvianolic acid B.

[0038] Example 2

[0039] Verification of in vitro 3D blood-brain barrier model

[0040] A 3D microsphere blood-brain barrier model was used to simulate ischemia-reperfusion injury and evaluate the effects of the traditional Chinese medicine combination on permeability, protein expression, and inflammatory pathways:

[0041] 1. Construction of 3D blood-brain barrier model: Human brain microvascular endothelial cells (HBMEC), human astrocytes (HA), and human brain vascular pericytes (HBVP) were co-cultured. Using EGM-2-MV medium (microvascular endothelial cell growth medium-2, add 25 mL of FBS, 0.2 mL of Hydrocortisone, 2 mL of hFGF-B, 0.5 mL of R3-IGF-1, 0.5 mL of Ascorbic Acid, 0.5 mL of hEGF, and 0.5 mL of GA-1000 to every 500 mL of EGM-2-MV), HBMEC, HBVP, and HA were inoculated into a low-attachment well plate at a ratio of 1500:1500:1500 and placed in a 37 °C constant temperature incubator containing 95% air and 5% CO2 for culture. After about 48 h, 3D blood-brain barrier microspheres were formed.

[0042] 2. Construction of ischemia-reperfusion model: 48 h after the formation of blood-brain barrier spheres, an oxygen-glucose deprivation / reoxygenation (oxygen-glucose deprivation / reoxygenation) model was established to simulate brain ischemia-reperfusion injury. The EGM-2-MV medium was replaced with glucose-free DMEM medium, and the culture environment was changed to a 37 °C incubator containing 95% N2 and 5% CO2 to achieve hypoxic conditions. After 6 h of oxygen-glucose deprivation, the glucose-free DMEM medium was replaced with EGM-2-MV medium, and reoxygenation culture was carried out in a 37 °C humidified incubator with 5% CO2 and 95% natural air for 12 h.

[0043] 3. Drug treatment: The blood-brain barrier spheroids were divided into a control group (anoxic glucose deprivation / reoxygenation) and an oxygen-glucose deprivation / reoxygenation group (the blood-brain barrier spheroids were treated with oxygen-glucose deprivation / reoxygenation); Salvia miltiorrhiza - Ligusticum chuanxiong (for the traditional Chinese medicine composition of the present invention, the blood-brain barrier spheroids (BBB spheroid) were cultured in EGM-2-MV containing 3.0 μL / mL of Salvia miltiorrhiza - Ligusticum chuanxiong, 12 h before oxygen-glucose deprivation and 6 h after oxygen-glucose deprivation); salvianolic acid B (the blood-brain barrier spheroids were cultured in EGM-2-MV containing 5.67 μM of salvianolic acid B for 12 h, with oxygen-glucose deprivation / reoxygenation); senkyunolide I (the blood-brain barrier spheroids were cultured in EGM-2-MV containing 1 μM of senkyunolide I for 12 h before oxygen-glucose deprivation / reoxygenation); 5.67 μM of salvianolic acid B and 1 μM of senkyunolide I were used to treat the 3D blood-brain barrier model alone or in combination, and the effects on blood-brain barrier permeability, cell viability, and tight junction protein expression were observed.

[0044] The permeability of the blood-brain barrier spheroids was evaluated by retaining extracellularly delivered FITC-labeled dextran. OGD / r-induced blood-brain barrier leakage increased FITC-labeled dextran (10KDa and 70KDa) to 48.5% and 29.9% respectively. Meanwhile, pretreatment with Salvia miltiorrhiza - Ligusticum chuanxiong or the positive control drug edaravone (EDA) prevented FITC-labeled dextran (70KDa and 10KDa) from entering the organoids, reducing the permeability to 74.8% and 67.6% respectively. The results are as Figure 2 shown, and both results show that Salvia miltiorrhiza - Ligusticum chuanxiong reduces the permeability of the blood-brain barrier.

[0045] Zonula occludens-1 is a core protein of tight junctions, which maintains the integrity of the intercellular barrier by connecting transmembrane proteins (such as claudin-5 and occludin) to the cytoskeleton. In the blood-brain barrier (BBB), the distribution and expression level of ZO-1 are important indicators for evaluating barrier function. The expression of zonula occludens-1 in Salvia miltiorrhiza - Ligusticum chuanxiong and the matrix metalloproteinase 9 inhibitor SB-3CT increased by 0.23 and 0.24 times, and that of occludin increased by 0.71 and 0.45 times. Importantly, the expression of zonula occludens-1 in salvianolic acid B and senkyunolide I increased by 0.13 and 0.07 times, and that of occludin increased by 0.53 and 0.40 times. Salvianolic acid B and senkyunolide I achieved better results than each alone, increasing the expression of zonula occludens- by 0.17 times and that of occludin by 0.62 times. The protective effects of salvianolic acid B, senkyunolide I, and their combination on the blood-brain barrier were evaluated after oxygen-glucose deprivation / reoxygenation. The results are as Figure 3 shown that the functional proteins of zonula occludens-1 and occludin decreased after oxygen-glucose deprivation / reoxygenation injury, while those of salvianolic acid B, senkyunolide I, salvianolic acid B - senkyunolide I, and SB-3CT increased.

[0046] Example 3

[0047] Molecular mechanism study

[0048] Combined with molecular docking, surface plasmon resonance and molecular dynamics simulation, clarify the interaction between salvianolic acid B and senkyunolide I and the target matrix metalloproteinase 9:

[0049] 1. IPA transcriptome core analysis: RNA-seq and network analysis showed that neuroinflammation mediated by the toll-like receptor 4-NF-κB-matrix metalloproteinase 9 signaling pathway is the key mechanism for the combination of Salvia miltiorrhiza and Ligusticum chuanxiong components to cause blood-brain barrier damage

[0050] The results are as Figure 4 shown. Through transcriptome sequencing and analysis, a total of 1,867 differentially expressed genes were identified in the Salvia miltiorrhiza-Ligusticum chuanxiong component combination group and the acute ischemic stroke group (1 day). In this follow-up study, 102 blood-brain barrier-related differentially expressed genes were extracted from 1,867 differentially expressed genes, of which 36 were up-regulated and 66 were down-regulated, with a cut-off value of fold change ≥ 2 and p value ≤ 0.05, and submitted for IPA analysis. The top 3 canonical signaling pathways with the highest correlation were the atherosclerotic signaling pathway, the neuroinflammatory signaling pathway, and the IL-15 production signaling pathway. The top 3 diseases and functions were intercellular signaling and interaction, nervous system diseases, and organism survival, as shown by the decreasing logarithm (p value) score. Further pathway interaction analysis showed that matrix metalloproteinase 9, an important protein that maintains blood-brain barrier integrity expressed in astrocytes and endothelial cells, was involved in 7 of the top 10 pathways. In addition, the relative p value of the toll-like receptor 4 gene was also ranked high among the top 10 upstream regulators. In addition, toll-like receptor 4-NF-κB-matrix metalloproteinase 9-mediated blood-brain barrier damage is the core part of the neuroinflammatory signaling pathway. In summary, the toll-like receptor 4-NF-κB-matrix metalloproteinase 9-mediated neuroinflammatory signaling pathway is an important potential pharmacological mechanism for the protection of the blood-brain barrier by Salvia miltiorrhiza and Ligusticum chuanxiong after acute ischemic stroke.

[0051] 2. Molecular docking: Using molecular docking software, study the binding ability of salvianolic acid B and senkyunolide I in the traditional Chinese medicine composition containing salvianolic acid B and senkyunolide I to matrix metalloproteinase 9.

[0052] Among them, 25 compounds with high affinity for matrix metalloproteinase 9 protein were screened out by molecular docking. Senkyunolide I from Salvia miltiorrhiza and Ligusticum chuanxiong are two representative potential active ingredients against matrix metalloproteinase 9. The two-dimensional and three-dimensional interaction diagrams of salvianolic acid B-matrix metalloproteinase 9 and senkyunolide I-matrix metalloproteinase 9 are as Figure 5 shown in Figure B.

[0053] 3. Surface Plasmon Resonance (SPR) experiment: Verify the binding affinity of salvianolic acid B and ligustilide I to matrix metalloproteinase 9 through surface plasmon resonance technology.

[0054] As Figure 5 shown, surface plasmon resonance was used to measure the component - matrix metalloproteinase 9 interaction, and the binding response in the resonance unit was continuously recorded in real time. Salvianolic acid B increased the surface plasmon resonance reaction in a dose - dependent manner, with a KD value of 4.90×10 - 7 μM. Dynamic simulation of salvianolic acid B - matrix metalloproteinase 9 was performed according to the reported method. The salvianolic acid B - matrix metalloproteinase 9 binding complex was stable, and after a rapid increase within the first 15 ns, the root - mean - square deviation (RMSD) value stabilized at The residues at the ligand - binding site showed sufficient stability, with low root - mean - square fluctuation (RMSF) values. Gibbs free energy showed that the binding energy of salvianolic acid B was approximately - 78 kJ / mol, indicating a strong binding effect between salvianolic acid B and matrix metalloproteinase 9.

[0055] 4. Molecular dynamics simulation: Study the binding stability of salvianolic acid B and ligustilide I to matrix metalloproteinase 9 through molecular dynamics simulation.

[0056] By analyzing the superimposed images of frames captured every 5 ns during the molecular dynamics (MD) simulation process, it was found that the binding of the ligand and the enzyme at the binding site was more stable. The results as Figure 5 Figure H shows that the 2 - (3′,4′ - dihydroxyphenyl) - benzofuran ring of the compound remained stable within 20 ns. This change can be attributed to the large and flexible structure of the ligand and its binding position on the enzyme. During the entire molecular dynamics simulation process, the salvianolic acid B - matrix metalloproteinase 9 complex formed an average of 4 - 5 hydrogen bonds, including the matrix metalloproteinase 9 residues of Arg51, Thr109, Asp177, Tyr179, Asp201, Gly100, Glu111, Gln108, Tyr50, Leu104, and Gly178. The hydrogen - bond occupancy of Arg51, Thr109, Asp177, and Tyr179 was between 58.07% and 21.78%, indicating a stable interaction between salvianolic acid B and matrix metalloproteinase 9.

[0057] 5. Detection of NF - κB - matrix metalloproteinase 9 axis proteins by 3D microspheres of the blood - brain barrier

[0058] As Figure 6The immunofluorescence analysis shown above confirmed that the TOLL-like receptor 4-NF-κB-matrix metalloproteinase 9-mediated neuroinflammatory signaling pathway is the key mechanism by which danshen and chuanxiong protect the blood-brain barrier. After oxygen-glucose deprivation / reoxygenation injury, the fluorescence intensities of TOLL-like receptor 4, matrix metalloproteinase 9, and NF-κB nuclear translocation increased by 44.1%, 27.6%, and 40.2%, respectively ( Figure 6 B). Similar to the TOLL-like receptor 4 inhibitor TAK-242, after administration, the fluorescence intensities of TOLL-like receptor 4 and matrix metalloproteinase 9 decreased by 22% and 10.3%, respectively. Danshen and chuanxiong reduced NF-κB nuclear translocation by 14.1%( Figure 6 A, Figure 6 B).

[0059] Example 4

[0060] Establishment of an in vivo mouse model of ischemic reperfusion injury and drug treatment

[0061] Using the mouse middle cerebral artery occlusion model, the protective effects of the combination on blood-brain barrier integrity, cerebral infarction volume, and neurological function were analyzed:

[0062] 1. Establishment of the mouse ischemic reperfusion injury model: By isolating the left common carotid artery, external carotid artery, internal carotid artery, and pterygopalatine artery of the mouse, a middle cerebral artery occlusion (MCAO) model was established to simulate ischemic stroke. The left common carotid artery, external carotid artery, internal carotid artery, and pterygopalatine artery were isolated to establish the middle cerebral artery occlusion model. Then, the external carotid artery was ligated with a medical suture, the external carotid artery was incised, and a silicone-coated nylon monofilament (L1800, Jialing Biotechnology Co., Ltd., Guangzhou) was inserted. The cerebral blood flow was measured immediately after 30 min of ischemia. When the monofilament was removed, the external carotid artery was ligated. Then, after monitoring the restoration of cerebral blood flow, the skin of the head and neck was sutured. The sham operation group had the same surgical procedure as the model group except that no monofilament was placed. A temperature-controlled heating pad was used to maintain the rectal temperature of the mouse at 37.0°C ± 0.5°C, and all physiological parameters were maintained within the normal range before and after the operation.

[0063] 2. Drug treatment: A traditional Chinese medicine injection containing salvianolic acid B and ligustilide I, as well as salvianolic acid B and ligustilide I alone or in combination, were used to treat the mice. After cerebral ischemia reperfusion injury, they were injected intravenously. The sham operation group and the model group were given an equal amount of normal saline.

[0064] 3. Result evaluation: The permeability of the blood-brain barrier was evaluated by Evans blue staining, the cerebral infarction volume was evaluated by TTC staining, and the neurological deficit was evaluated by neurological function scoring.

[0065] The results are as Figure 7As shown, in a mouse model of 3-day cerebral ischemia-reperfusion injury, the integrity of the blood-brain barrier was evaluated by the Evans blue extravasation method. Danshen-Chuanxiong (2.5 mL / kg, 5 mL / kg, and 10 mL / kg, 0.65 - 1.20 times) or edaravone (5 mL / kg, 0.70 times) significantly reduced Evans blue leakage. The infarct volume in the cerebral ischemia-reperfusion injury group was 36.8% as determined by TTC staining. After treatment with Danshen-Chuanxiong and edaravone, neurological deficits were significantly reduced, to 23.9% and 20.7% respectively, and at the same time, neurological deficits were significantly alleviated.

[0066] 4. Mechanism evaluation: Western blotting and immunohistofluorescence were used to detect the protein expression of the Toll-like receptor 4-NF-κB-matrix metalloproteinase 9 axis.

[0067] As Figure 8 shown, Western blotting and immunohistofluorescence were used to detect the protein of the Toll-like receptor 4-NF-κB-matrix metalloproteinase 9 axis. The results were as Figure 8 shown in A - D. The expressions of Toll-like receptor 4 (0.41 times), p-NF-κB (1.33 times), and matrix metalloproteinase 9 (0.41 times) in the brain tissues of CIRI mice were significantly increased after cerebral ischemia-reperfusion injury. Danshen-Chuanxiong decreased their expressions by 0.39, 0.45, and 0.36 times respectively ( Figure 8 A - D). Similarly, IF significantly increased the expressions of Toll-like receptor 4 (0.77 times), p-NF-κB (0.62 times), and matrix metalloproteinase 9 (1.25 times). Danshen-Chuanxiong decreased the expressions of Toll-like receptor 4, p-NF-κB, and matrix metalloproteinase 9 by 0.37, 0.29, and 0.37 times respectively. Compared with the cerebral ischemia-reperfusion injury group, the expressions of ZO-1 tight junction protein (1.66 times) and occludin tight junction protein (1.61 times) in Danshen-Chuanxiong were increased (1.69 times and 1.64 times). It can be seen that the traditional Chinese medicine composition described in the present invention maintains the integrity of the blood-brain barrier by regulating the Toll-like receptor 4 / NF-κB / matrix metalloproteinase 9-mediated neuroinflammatory signaling pathway and plays a protective role in ischemic stroke in 3D spheres and mice.

[0068] In summary, the traditional Chinese medicine composition described in the present invention was verified by an in vitro 3D blood-brain barrier model and an in vivo mouse ischemia-reperfusion model. It can significantly reduce the increase in blood-brain barrier permeability caused by ischemia-reperfusion injury, maintain the expression levels of tight junction proteins ZO-1 and Occludin, and inhibit the activation of the Toll-like receptor 4 / NF-κB / matrix metalloproteinase 9 neuroinflammatory signaling pathway. This combination enhances the protective effect of the blood-brain barrier through a synergistic effect, provides a new strategy for the treatment of ischemic stroke, and has significant clinical application value.

[0069] The above-described embodiments are merely descriptions of the preferred embodiments of the present invention and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations and improvements made by those of ordinary skill in the art to the technical solutions of the present invention shall fall within the protection scope of the present invention.

Claims

1. Use of a traditional Chinese medicine composition in the preparation of a blood-brain barrier protector, characterized in that: The traditional Chinese medicine composition is composed of salvianolic acid B and senkyunolide I.

2. The use according to claim 1, wherein: The weight ratio of salvianolic acid B to senkyunolide I is 1:2.6 - 3.

3. The use according to claim 1, characterized in that: The cell dosages of salvianolic acid B and senkyunolide I are 5.7 μMole and 1 μMole respectively.

4. The use according to any one of claims 1 to 3, characterized in that: The blood - brain barrier protector is a drug for preventing or treating ischemic stroke.

5. The use according to claim 1, wherein: The traditional Chinese medicine composition is prepared by the following method: (1) Weigh the monomer standard products of senkyunolide I and salvianolic acid B respectively according to the amounts; (2) After dissolving the monomer standard products of senkyunolide I and salvianolic acid B in methanol respectively, dilute and mix them with pure water.

6. A blood-brain barrier protectant, characterized in that: It includes the traditional Chinese medicine composition described in claim 1 and pharmaceutically acceptable excipients.

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